scholarly journals Sen1 is required for RNA Polymerase III transcription termination in an R-loop independent manner

2019 ◽  
Author(s):  
Julieta Rivosecchi ◽  
Marc Larochelle ◽  
Camille Teste ◽  
Frédéric Grenier ◽  
Amélie Malapert ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Fission Yeast ◽  
Rna Polymerase Ii ◽  
Genome Stability ◽  
Transcription Termination ◽  
Rna Polymerase Iii ◽  
Rna Polymerases ◽  
Independent Manner ◽  
Polymerase Iii

ABSTRACTR-loop disassembly by the human helicase Senataxin contributes to genome stability and to proper transcription termination at a subset of RNA polymerase II genes. Whether Senataxin-mediated R-loop disassembly also contributes to transcription termination at other classes of genes has remained unclear. Here we show in fission yeast that SenataxinSen1promotes efficient termination of RNA Polymerase III (RNAP3) transcriptionin vivo. In the absence of SenataxinSen1, RNAP3 accumulates downstream of the primary terminator at RNAP3-transcribed genes and produces long exosome-sensitive 3’-extended transcripts. Importantly, neither of these defects was affected by the removal of R-loops. The finding that SenataxinSen1acts as an ancillary factor for RNAP3 transcription terminationin vivochallenges the pre-existing view that RNAP3 terminates transcription autonomously. We propose that Senataxin is a cofactor for transcription termination that has been co-opted by different RNA polymerases in the course of evolution.

scholarly journals Termination-altering mutations in the second-largest subunit of yeast RNA polymerase III.

1995 ◽  
Vol 15 (3) ◽  
pp. 1467-1478 ◽  
Author(s):  
S A Shaaban ◽  
B M Krupp ◽  
B D Hall
Keyword(s):  
Amino Acid ◽  
Rna Polymerase ◽  
Transcription Initiation ◽  
Transcription Termination ◽  
Rna Polymerase Iii ◽  
In Vitro Transcription ◽  
The Other ◽  
Polymerase Iii

In order to identify catalytically important amino acid changes within the second-largest subunit of yeast RNA polymerase III, we mutagenized selected regions of its gene (RET1) and devised in vivo assays for both increased and decreased transcription termination by this enzyme. Using as the reporter gene a mutant SUP4-o tRNA gene that in one case terminates prematurely and in the other case fails to terminate, we screened mutagenized RET1 libraries for reduced and increased transcription termination, respectively. The gain in suppression phenotype was in both cases scored as a reduction in the accumulation of red pigment in yeast strains harboring the ade2-1 ochre mutation. Termination-altering mutations were obtained in regions of the RET1 gene encoding amino acids 300 to 325, 455 to 486, 487 to 521, and 1061 to 1082 of the protein. In degree of amino acid sequence conservation, these range from highly variable in the first to highly conserved in the last two regions. Residues 300 to 325 yielded mainly reduced-termination mutants, while in region 1061 to 1082, increased-termination mutants were obtained exclusively. All mutants recovered, while causing gain of suppression with one SUP4 allele, brought about a reduction in suppression with the other allele, thus confirming that the phenotype is due to altered termination rather than an elevated level of transcription initiation. In vitro transcription reactions performed with extracts from several strong mutants demonstrated that the mutant polymerases respond to RNA terminator sequences in a manner that matches their in vivo termination phenotypes.

Intrinsic sites of transcription termination and pausing in the c-myc gene

1988 ◽  
Vol 8 (10) ◽  
pp. 4389-4394
Author(s):  
T K Kerppola ◽  
C M Kane
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Xenopus Oocytes ◽  
Transcription Termination ◽  
Transcription Elongation ◽  
Rna Polymerase Iii ◽  
Intron 1 ◽  
Exon 1 ◽  
Myc Gene

We have studied transcription elongation and termination in the human c-myc gene. Transcription of c-myc gene sequences with purified mammalian RNA polymerase II revealed several sites of transcription termination and pausing in the vicinity of the exon 1-intron 1 junction. This region previously has been shown to block transcription elongation in vivo by nuclear run-on analysis (D. Bentley and M. Groudine, Nature [London] 321:702-706, 1986). These sites were recognized by purified RNA polymerase II, and we therefore designated them intrinsic sites of termination and pausing. Two of these sites cause termination of RNA polymerase III transcription as well. RNA polymerase II terminated transcription in a cluster of seven consecutive T residues in the nontranscribed strand and paused during transcription at three additional sites in this region. The intrinsic sites of transcription termination and pausing described here correspond closely to the 3' ends of transcripts synthesized in Xenopus oocytes injected with plasmids containing the c-myc termination region (D. Bentley and M. Groudine, Cell 53:245-256, 1988). This correspondence suggests that the intrinsic recognition of these termination and pause sites by purified RNA polymerase II may play a role in the transcription elongation block observed in vivo.

Specificity of RNA maturation pathways: RNAs transcribed by RNA polymerase III are not substrates for splicing or polyadenylation

1987 ◽  
Vol 7 (10) ◽  
pp. 3602-3612
Author(s):  
S S Sisodia ◽  
B Sollner-Webb ◽  
D W Cleveland
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Rna Polymerase Iii ◽  
Rnase H ◽  
Intact Cells ◽  
S1 Nuclease ◽  
Polymerase Iii ◽  
Hybrid Genes ◽  
Rna Maturation

To analyze the specificity of RNA processing reactions, we constructed hybrid genes containing RNA polymerase III promoters fused to sequences that are normally transcribed by polymerase II and assessed their transcripts following transfection into human 293 cells. Transcripts derived from these chimeric constructs were analyzed by using a combined RNase H and S1 nuclease assay to test whether RNAs containing consensus 5' and 3' splicing signals could be efficiently spliced in intact cells, even though they were transcribed by RNA polymerase III. We found that polymerase III-derived RNAs are not substrates for splicing. Similarly, we were not able to detect poly(A)+ RNAs derived from genes that contained a polymerase III promoter linked to sequences that were necessary and sufficient to direct 3'-end cleavage and polyadenylation when transcribed by RNA polymerase II. Our findings are consistent with the view that in vivo splicing and polyadenylation pathways are obligatorily coupled to transcription by RNA polymerase II.

scholarly journals Transcription Termination by RNA Polymerase III in Fission Yeast

2000 ◽  
Vol 275 (37) ◽  
pp. 29076-29081 ◽  
Author(s):  
Mitsuhiro Hamada ◽  
Amy L. Sakulich ◽  
Shashi B. Koduru ◽  
Richard J. Maraia
Keyword(s):  
Rna Polymerase ◽  
Fission Yeast ◽  
Transcription Termination ◽  
Rna Polymerase Iii ◽  
Polymerase Iii

scholarly journals Intrinsic sites of transcription termination and pausing in the c-myc gene.

1988 ◽  
Vol 8 (10) ◽  
pp. 4389-4394 ◽  
Author(s):  
T K Kerppola ◽  
C M Kane
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Xenopus Oocytes ◽  
Transcription Termination ◽  
Transcription Elongation ◽  
Rna Polymerase Iii ◽  
Intron 1 ◽  
Exon 1 ◽  
Myc Gene

We have studied transcription elongation and termination in the human c-myc gene. Transcription of c-myc gene sequences with purified mammalian RNA polymerase II revealed several sites of transcription termination and pausing in the vicinity of the exon 1-intron 1 junction. This region previously has been shown to block transcription elongation in vivo by nuclear run-on analysis (D. Bentley and M. Groudine, Nature [London] 321:702-706, 1986). These sites were recognized by purified RNA polymerase II, and we therefore designated them intrinsic sites of termination and pausing. Two of these sites cause termination of RNA polymerase III transcription as well. RNA polymerase II terminated transcription in a cluster of seven consecutive T residues in the nontranscribed strand and paused during transcription at three additional sites in this region. The intrinsic sites of transcription termination and pausing described here correspond closely to the 3' ends of transcripts synthesized in Xenopus oocytes injected with plasmids containing the c-myc termination region (D. Bentley and M. Groudine, Cell 53:245-256, 1988). This correspondence suggests that the intrinsic recognition of these termination and pause sites by purified RNA polymerase II may play a role in the transcription elongation block observed in vivo.

scholarly journals ret1-1, a yeast mutant affecting transcription termination by RNA polymerase III.

Genetics ◽  
1990 ◽  
Vol 125 (2) ◽  
pp. 293-303
Author(s):  
P James ◽  
B D Hall
Keyword(s):  
Rna Polymerase ◽  
Transcription Termination ◽  
Rna Polymerase Iii ◽  
In Vitro Transcription ◽  
Polymerase Iii ◽  
Reconstituted System ◽  
Gene Fractionation ◽  
Yeast Mutants

Abstract In eukaryotes, extended tracts of T residues are known to signal the termination of RNA polymerase III transcription. However, it is not understood how the transcription complex interacts with this signal. We have developed a selection system in yeast that uses ochre suppressors weakened by altered transcription termination signals to identify mutations in the proteins involved in termination of transcription by RNA polymerase III. Over 7600 suppression-plus yeast mutants were selected and screened, leading to the identification of one whose effect is mediated transcriptionally. The ret1-1 mutation arose in conjunction with multiple rare events, including uninduced sporulation, gene amplification, and mutation. In vitro transcription extracts from ret1-1 cells terminate less efficiently at weak transcription termination signals than those from RET1 cells, using a variety of tRNA templates. In vivo this reduced termination efficiency can lead to either an increase or a further decrease in suppressor strength, depending on the location of the altered termination signal present in the suppressor tRNA gene. Fractionation of in vitro transcription extracts and purification of RNA polymerase III has shown that the mutant effect is mediated by highly purified polymerase in a reconstituted system.

scholarly journals Specificity of RNA maturation pathways: RNAs transcribed by RNA polymerase III are not substrates for splicing or polyadenylation.

1987 ◽  
Vol 7 (10) ◽  
pp. 3602-3612 ◽  
Author(s):  
S S Sisodia ◽  
B Sollner-Webb ◽  
D W Cleveland
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Rna Polymerase Iii ◽  
Rnase H ◽  
Intact Cells ◽  
S1 Nuclease ◽  
Polymerase Iii ◽  
Hybrid Genes ◽  
Rna Maturation

To analyze the specificity of RNA processing reactions, we constructed hybrid genes containing RNA polymerase III promoters fused to sequences that are normally transcribed by polymerase II and assessed their transcripts following transfection into human 293 cells. Transcripts derived from these chimeric constructs were analyzed by using a combined RNase H and S1 nuclease assay to test whether RNAs containing consensus 5' and 3' splicing signals could be efficiently spliced in intact cells, even though they were transcribed by RNA polymerase III. We found that polymerase III-derived RNAs are not substrates for splicing. Similarly, we were not able to detect poly(A)+ RNAs derived from genes that contained a polymerase III promoter linked to sequences that were necessary and sufficient to direct 3'-end cleavage and polyadenylation when transcribed by RNA polymerase II. Our findings are consistent with the view that in vivo splicing and polyadenylation pathways are obligatorily coupled to transcription by RNA polymerase II.

scholarly journals Template Structural Requirements for Transcription In Vivo by RNA Polymerase II

1982 ◽  
Vol 2 (12) ◽  
pp. 1595-1607 ◽  
Author(s):  
Timothy J. Miller ◽  
Janet E. Mertz
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Rna Polymerase Iii ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
Structural Requirements ◽  
Infected Monkey ◽  
Full Complement ◽  
Sv40 Dna

Purified simian virus 40 (SV40) DNA is reconstituted into chromatin and transcribed by endogenous RNA polymerase II when microinjected into nuclei ofXenopus laevisoocytes. We have correlated the kinetics of chromatin reconstitution with that of accumulation of virus-specific RNA in this system. A delay of approximately 3 h was found in the appearance of appreciable numbers of both fully supercoiled molecules and transcriptionally active templates. SV40 minichromosomes, isolated from virus-infected monkey cells with 0.2 M NaCl, also exhibited this lag in onset of transcriptional activity when microinjected into oocytes. These findings indicate that neither purified SV40 DNA nor SV40 DNA containing a full complement of nucleosomes can function as a template for transcription in vivo before association with appropriate cellular nonhistone chromosomal factors has taken place. In addition, the gradual degradation of linear SV40 DNA in oocytes was not sufficient to account for the fact that it was much less transcriptionally active than circular SV40 DNA. Taken together, these results indicate that the conformational state of the DNA can affect its ability to function as a template for transcription in vivo by RNA polymerase II. In contrast, transcription by RNA polymerase III of purified, circularized cloned DNAs encoding genes for 5S rRNA was detectable long before the injected DNAs had time to reconstitute into chromatin. Therefore, the template structural requirements for transcription in vivo by RNA polymerases II and III are different.

A common octamer motif binding protein is involved in the transcription of U6 snRNA by RNA polymerase III and U2 snRNA by RNA polymerase II

Cell ◽  
1987 ◽  
Vol 51 (1) ◽  
pp. 71-79 ◽  
Author(s):  
Philippe Carbon ◽  
Sylvie Murgo ◽  
Jean-Pierre Ebel ◽  
Alain Krol ◽  
Graham Tebb ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Binding Protein ◽  
Rna Polymerase Iii ◽  
U2 Snrna ◽  
U6 Snrna ◽  
Polymerase Iii ◽  
Octamer Motif
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